Genome-wide (ChIP-seq) Identification of Target Genes Regulated by WRKY33 During Submergence Stress in Arabidopsis

Abstract Background: Hypoxia induced by flooding causes significant losses to crop production almost every year. However, the molecular network of hypoxia signaling pathway is still poorly understood. According to previous studies, transgenic plants overexpressing the WRKY33 gene showed enhanced resistance to submergence stress. Thus, the transcription factor WRKY33 may regulate a series of target genes in response to submergence. Here, to determine the putative downstream targets of WRKY33 at a genome-wide scale in Arabidopsis thaliana, we performed the chromatin immunoprecipitation sequencing (ChIP-seq) using 35S:FLAG-WRKY33 overexpression transgenic lines (WRKY33-OE) upon 24 hours of submergence treatment.Results: Using ChIP-seq data, we identified a total of 104 WRKY33-binding genes under submergence treatment (WRKY33BGHs). Most of the WRKY33BGHs are involved in the oxidation-reduction process, programmed cell death in response to reactive oxygen species, lipid biosynthesis process, and other processes related to stress responses. Moreover, the major motif identified in the WRKY33BGHs promoters is a new cis-element, TCTCTC (we named it as “TC box”). This cis-element is different from the previously known W box for WRKY33. Further qPCR experiments verified that genes carrying this motif in their promoters could be regulated by WRKY33 upon submergence treatment.Conclusions: Our study has identified a new putative binding motif of WRKY33 and recovered numerous previously unknown target genes of WRKY33 during submergence stress. The WRKY33 gene positively participates in flooding response probably by transcriptional regulation of the downstream hypoxia-related target genes via a “TC box”.